Asthma is an allergic inflammatory response to environmental challenges (e.g., house dust mite (HDM) or pollen). These sources of environmental antigens commonly trigger type 2 inflammation. This proposal seeks to identify a very novel mechanism coordinating the immune system via a thrombocyte (platelet)-derived factor to orchestrate type 2 inflammation. In our preliminary studies, the Wnt antagonist Dickkopf-1 (Dkk-1) is necessary for a pathological response to intranasal challenge with HDM extract. Interestingly, our preliminary results suggest that the primary source of this Dkk-1 is from platelets and activation of platelets results in stimulation of Dkk-1 release. The activated platelets can then physically bind neutrophils which facilitates migration to the lung tissue by forming leukocyte-platelet aggregates (LPA). Other investigators have shown that thymic stromal lymphopoietin (TSLP) is also critical for initiating the type 2 immune response. We observed that the elevation of circulating Dkk-1 levels in response to HDM can also be blocked with TSLP neutralization, suggesting that the environmental allergen can trigger TSLP release from epithelial cells, and then TSLP activates platelets via TSLP-R. We hypothesize that Dkk-1 utilizes unique signaling molecules that define the Th2 differentiation pathway in CD4 T cells. In Aim 1, the platelet contribution will be tested using mice conditionally deleted for Dkk-1 or TSLP-R in platelets. Use of these two strains will address the hypothesis that TSLP is an important stimulatory ligand to release Dkk-1 from platelets. In Aim 2 we will determine whether LRP6 in CD4 T cells is critical for this response. We will evaluate the type 2 immune responses in these strains both in vivo and in vitro in the HDM-induced asthma model. We will also test the ability of purified HDM derived proteases can activate platelets and if antibody to Dkk-1 can block the response. In addition to the standard analysis of this model, we will analyze the formation of LPA, and will further characterize the effects on cell migration into the lung tissue. We anticipate that this study will identify new immunological mechanisms that are critical to the induction and maintenance of chronic type 2 immune responses. Since TSLP and Th2 cell differentiation and type 2 cytokine production are commonly found in other type 2 immune responses such as parasite infections or skin inflammation (e.g. atopic dermatitis) our results will be important for type 2 inflammatory diseases in general.